Genetic control elements in herpes simplex virus type 1 (HSV-1) are investigated. Deletions occuring in a 9.5 base pair EcoR1 defective DNA fragment co-transfected with the HSV-1 tk gene in mouse Ltk- cells are analyzed. Tumors generated in vivo from an inoculation of cells transformed in vitro by HSV-1 were negative for retention of virus-specific sequences.